二元官能基自組裝單層膜修飾表面之生物應用

Biological Application of Surfaces Tailored with Binary Self-Assembled Monolayers (SAMs)

張薰勻(Hsun-Yun Chang)

指導教授 ： 薛景中 ； 曾繁根

國立清華大學/原子科學院/工程與系統科學系/博士(2015年)

摘要

摘要
自組裝單層膜為改變表面性質的強大技術，且廣泛地應用在學術研究與工業應用．材料的選擇無論是塊材還是奈米粒子，自組裝單層摩的組成提供廣泛的選擇以利於實驗的要求．對於生物的應用上，硫醇修飾金基材的自組裝單層膜系統被廣泛的使用來發展生物晶片、生物感測器、基因傳輸者等研究。於是研究細胞與有著特殊表面性質的自組裝單層膜修飾之金材料之間的交互作用為有其必要。隨著二元官能基自組裝單層膜修飾表面之金材料的發展，細胞與修飾過的材料表面之交互作用得以更進一步的研究。在此研究當中，帶有一系列不同表面電位的二元官能基自組裝單層膜修飾表面之金奈米粒子與金表面塊材均製備出來以進行帶有不同表面電位的二元官能基自組裝單層膜修飾表面之金材料對於生物應用的研究。

在金奈米粒子的生物應用部分，研究結果發現帶有不同表面電位的電位的二元官能基自組裝單層膜修飾表面之金奈米粒子對細胞不帶有毒性，且能被細胞所攝取．金奈米粒子於細胞內部的影像使用掃描透射電子顯微鏡呈現。細胞對於金奈米粒子的攝取量與金奈米粒子的表面電位為正相關。被攝取後的金奈米粒子在細胞中的位置將進一步的利用飛行時間二次離子質譜儀研究．而對於帶有不同表面電位的金奈米粒子作為運送者來傳送帶有表面螢光蛋白質的質體的能力，研究結果發現金奈米粒子的表面電位搭配細胞包吞作用過程中的酸鹼值會影響到運送的結果，帶有特定表面電位的金奈米粒子能在核內體保護質體不被降解並在包吞作用後於細胞質中釋出質體使其作用，而發揮最大的傳送能力。

另外，纖維母細胞與上皮細胞於二元官能基自組裝單層膜修飾表面之金基材的細胞行為也進行研究，包含細胞的貼附、增殖與形態。實驗結果發現，隨著材料的表面電位增加，纖維母細胞與上皮細胞的貼附數量均增加，且上皮細胞的貼附數量比纖維母細胞更為顯著。而表面電位對於兩種細胞的增殖行為並沒有影響。而對於細胞表面形貌，表面電位不會使上皮細胞的形貌改變，而會改變纖維母細胞的形貌。表面電位對於細胞種類間的差異可從細胞外間質主要的蛋白質組成解釋，相較於黏連蛋白為細胞外間質主要蛋白的纖維母細胞，層黏蛋白為細胞外間質主要蛋白的上皮細胞表現較高的貼附數量與不變的細胞形貌，這是由於帶正電的層黏蛋白於材料表面上的吸附量較高以利於上皮細胞的貼附。相較之下，由於帶負電的黏連蛋白在材料表面上的吸附量較少，使得以黏連蛋白作為主要細胞外間質蛋白的纖維母細胞必須改變其形貌來迎合不均勻的黏連蛋白區域之表面。

關鍵字：自組裝單層膜、表面電位、金、轉殖、細胞貼附

關鍵字

自組裝單層膜 ； 表面電位 ； 金 ； 基因轉殖 ； 細胞貼附

並列摘要

Being a powerful technique of modifying the surface property, Self-assembled monolayers (SAMs) has been employed for academia research and industrial application. The substrate can be bulk or nanoparticles; the SAMs composition provides wild selection for specific requirement. For biological application, the SAM system of thiols on Au susbtrate is of great interest on the development of biochip, biosensor, gene carriers, etc. Studying the interaction between cells and SAM-modified Au substrate with specific surface property has gained attention. With the development of binary SAM-modified Au substrates, the interaction between cell and SAM-modified Au substrates could be studied. In this work, binary SAM-modified Au substrates with a series of surface potential were made, including Au nanoparticles and bulks with Au surface. It was found that binary SAM-modified AuNPs have low cytotoxicity and can be internalized by cells that confirmed by STEM images, and the internalized AuNPs increased with increasing surface potential of AuNPs. The cellular distribution would be further studied using time-of-flight secondary ion mass spectroscopy (ToF-SIMS). For the capability of binary SAM–modified AuNPs as carriers to deliver DNA for expression, it was found to be relevant to the surface potential of AuNPs with respect to the change of pH value during delivery process. The highest delivery happened when AuNPs with moderate surface potential that can adsorb and protect DNA in endosome and release DNA in cytoplasm.

Moreover, cell behavior of fibroblasts and epithelial cells on binary SAM-modified Au surfaces were examined, including adhesion, proliferation and morphology. The results showed that the adhesion density of epithelial cells increased with increasing surface potential, which is similar to but varied more significantly compared with fibroblasts. The proliferation rate is found to be independent of surface potential in both cell types. Furthermore, epithelial cells show no morphological change with respect to surface potential, whereas the morphology of the fibroblasts clearly changed with the surface potential. These differences between the cell types were rationalized by considering the difference in ECM composition. Laminin-dominant epithelial cells showed higher adhesion density and less morphological change than did fibronectin-dominant fibroblasts because the more significant adsorption of positively charged laminin on the surface enhanced the adhesion of epithelial cells. In contrast, due to the dominance of negatively charged fibronectin that adsorbed weakly on the surface, fibroblasts had to change their morphology to fit the inhomogeneous fibronectin-adsorbed area.

並列關鍵字

self-assembled monolayers ； surface potential ； Au ； transfection ； cell adhesion

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